Umbilical cord blood (UCB) provides unique advantages over bone marrow (BM) and mobilized peripheral blood (MPB) as a source of hematopoietic stem cells (HSCs) for transplants, as it allows use of partially HLA-matched units without increasing GvHD incidence. Although UCB contains a higher frequency of primitive HSCs than BM and MPB, it contains significantly fewer total HSCs, which has severely limited its use in adult patients. Arteriocyte plans to address this unmet medical need with rapid, high yield ex vivo expansion of UCB cells using our NANEXTM expansion platform. The NANEXTM platform is based on a nano-fiber scaffold that mimics the bone marrow stem cell niche and has been shown to expand UCB total nucleated cells (TNCs) an average of 500-fold and CD34+ cells and average of 150-fold in 10-day culture. NANEXTM is currently available commercially as a Research Use Only product, but ultimately, Arteriocyte aims to develop this platform as a clinical product for use in umbilical cord blood transplant (UCBT). The objective of this Phase I proposal is to investigate the feasibility of using Arteriocyte's NANEXTM scaffold to expand UCB cells ex vivo as a potential means of overcoming the limited cell supply available from UCB for transplantation. We hypothesize that our NANEXTM scaffold will enable efficient production of high doses of UCB cells capable of reconstituting hematopoietic functions in an immune-compromised mouse model. This hypothesis will be tested in the following specific aims: Specific Aim 1: Optimization of Ex Vivo Expansion of UCB Cells on NANEXTM Ex vivo expansion of UCB cells on NANEXTM will be optimized by varying phenotype of the starting cell population, initial cell seeding density, cytokines, and culture time, and characterizing the frequency of CD34+ cells, colony forming cells (CFCs), and long-term culture-initiating cells (LTC-ICs). The optimized NANEXTM-expansion protocol will then be validated in a laboratory with significant experience with HSC transplantation at Dana-Farber Cancer Institute. Specific Aim 2: Evaluation of Engraftment of NANEXTM-Expanded UCB Cells in NOD/SCID/ 2microglobulin null Mice NANEX- and TCPS-expanded and non-expanded cells will be transplanted in a NOD/SCD/2microglobulinnull mouse model and the level of engraftment for each test groups will be evaluated using immunophenotyping/flow cytometry and Southern blotting (if necessary). The goal is to demonstrate that NANEX culture significantly increases the number of SCID repopulating cells (SRCs) compared to controls. If feasibility is demonstrated in this Phase I project, Arteriocyte will pursue further development of the NANEXTM system and evaluation in a clinical setting. For this reason, a team of collaborators has been assembled with significant experience in both pre-clinical and clinical UCBT, and the intention is to work together beyond this project. PUBLIC HEALTH RELEVANCE: Arteriocyte proposes to use our novel NANEXTM ex vivo expansion technology for the rapid, high yield production of cord blood-derived cells capable of long-term repopulation in vivo. We aim to test the feasibility of our approach by grafting NANEXTM-expanded cord blood cells in an immune-compromised mouse model. With further development, the NANEXTM system could enable the broader use of cord blood for restoring normal hematopoiesis in adult patients with hematologic cancers, anemias and a variety of other malignant and non- malignant disorders.